Bacteria that grow and reproduce solely inside the cells of other organisms exhibit a consistent pattern of genome reduction. In some cases, the reductive process is so extreme that the bacteria lack many genes thought to be essential for cellular life. Although it is generally assumed that these bacteria somehow rely on the genetic machinery of their host cell to offset the limitations of their degenerated genomes, the actual mechanistic basis for these intimate relationships between prokaryotic and eukaryotic genomes remains unclear. The proposed research will take advantage of recent advances in high throughput DNA sequencing technology to characterize the genomic interactions between psyllids (Insecta: Hemiptera) and their nutritional symbiont Carsonella, which exhibits one of the most extreme cases of bacterial genome reduction and degradation ever identified. The primary goal of this research is to determine the role of the host genome in enabling the bacterial symbionts to survive and reproduce in spite of their severely reduced gene content. The genomes from 7 Carsonella lineages and a related outgroup species will be sequenced and analyzed to comprehensively determine the functional capabilities retained by these bacteria. In addition, high throughput cDNA sequencing will be used to perform a genome-wide analysis of host gene expression. The bacteria are restricted to the cells of a specific organ in the insect host. Therefore, host gene expression in bacteria-containing cells will be compared to expression patterns in the rest of the body to identify host genes that are specifically upregulated in association with Carsonella. Functional annotation and analysis will be used to assess whether the identified host genes are capable of replacing key pathways that are no longer encoded within the symbiont genome. Furthermore, phylogenetic analysis will identify the origin of the host genes to determine whether they were co-opted from pre-existing host functions or acquired from a foreign source such as Carsonella or another bacterial lineage. This work will inform our understanding of the limits of bacterial evolution (i.e., "the minimal cell") and determine how interactions with eukaryotic genomes may allow intracellular bacteria to break through these limits. The results of this study will also be relevant to understanding the genomic function of the large number of intracellular bacteria that act as human pathogens. PUBLIC HEALTH RELEVANCE: Bacterial pathogens that live inside human cells have highly reduced genomes. Therefore, by identifying the mechanisms of bacterial function in the most extreme cases of genome degradation found in nature (nutritional bacteria living inside the cells of insects), this research could provide novel insights into the basic biology of infectious disease in humans.